Ten tips on immunosuppression in primary membranous nephropathy

ABSTRACT Membranous nephropathy (MN) management poses challenges, particularly in selecting appropriate immunosuppressive treatments (IST) and monitoring disease progression and complications. This article highlights 10 key tips for the management of primary MN based on current evidence and clinical experience. First, we advise against prescribing IST to patients without nephrotic syndrome (NS), emphasizing the need for close monitoring of disease progression. Second, we recommend initiating IST in patients with persistent NS or declining kidney function. Third, we suggest prescribing rituximab (RTX) or RTX combined with calcineurin inhibitors in medium-risk patients. Fourth, we propose cyclophosphamide-based immunosuppression for high-risk patients. Fifth, we discourage the use of glucocorticoid monotherapy or mycophenolate mofetil as initial treatments. Sixth, we underscore the importance of preventing infectious complications in patients receiving IST. Seventh, we emphasize the need for personalized monitoring of IST by closely measuring kidney function, proteinuria, serum albumin and anti-M-type phospholipase A2 receptor levels. Eighth, we recommend a stepwise approach in the treatment of resistant disease. Ninth, we advise adjusting treatment for relapses based on individual risk profiles. Finally, we caution about the potential recurrence of MN after kidney transplantation and suggest appropriate monitoring and treatment strategies for post-transplantation MN. These tips provide comprehensive guidance for clinicians managing MN, aiming to optimize patient outcomes and minimize complications.


TIP 1. DO NOT PRESCRIBE IMMUNOSUPPRESSIVE TREATMENT ( IST) TO PATIENTS WITHOUT NEPHROTIC SYNDROME ( NS)
Available data show that a high percentage of membranous nephropathy ( MN) cases with non-nephrotic proteinuria ( up to 40%) do not progress to nephrotic syndrome ( NS) [1 ].These cases should be closely followed without starting IST and watching the trend of the disease towards spontaneous complete remission, maintenance of non-nephrotic proteinuria or development of full NS.
Supporting a conservative attitude in these patients is a better tolerance to the amount of proteinuria in MN compared with other glomerular diseases such as immunoglobulin A nephropathy [2 ].On the other hand, the number of non-immunosuppressive antiproteinuric drugs has increased in recent years.Two large randomized trials [DAPA-CKD ( NCT03036150) and EMPA-KIDNEY ( NCT03594110) ] found that dapagliflozin and empagliflozin decrease the risk of CKD progression by 32% regardless of the presence of diabetes [3 -5 ].Both trials enrolled a combined total of 136 patients with MN [6 ].In a recent large retrospective cohort study, the use of sodium-glucose co-transporter 2 inhibitors ( SGLT2is) was associated with a proteinuria reduction of 48% after 1 year under SGLT2i in a diverse subset of primary and secondary glomerular diseases, including 89 patients with MN [7 ]. Results were similar irrespective of the underlying disease.Therefore, the combination of SGLT2i and renin-angiotensinaldosterone system ( RAAS) blockade can substantially reduce the amount of proteinuria in MN patients without complete NS.
An important limitation of the above statements is that the main studies on spontaneous remission of MN and on the natural history of MN with non-nephrotic proteinuria were published before the discovery of anti-M-type phospholipase A2 receptor ( PLA2R) antibodies and other MN-associated antigens [1 , 8 -10 ].Initial anti-PLA2R titres, and especially its evolution, should be carefully monitored in these patients [11 -13 ].

TIP 2. PRESCRIBE IST TO THOSE PATIENTS IN WHOM NS DOES NOT IMPROVE AFTER AN OBSERVATION PERIOD AND IN THOSE WITH DECLINING KIDNEY FUNCTION OR SERIOUS COMPLICATIONS OF NS
Spontaneous remission ( SR) is relatively frequent in MN ( ≈30% of cases) , especially in those with lower proteinuria [8 , 10 , 14 , 15 ].As well as the amount of proteinuria, other predictors of a higher probability of SR include female sex, baseline kidney function, treatment with RAAS blockers and a decrease in proteinuria > 50% in the first year.SR usually develops within 24 months since MN diagnosis and is generally associated with good long-term prognosis and low recurrence rate [10 ].Thus an observation period is highly recommended before IST is started.
In anti-PLA2R-positive cases, both baseline titres and the trajectory of the autoantibodies are crucial to determine the prognosis and the need for IST.Immunologic remission has been associated with clinical remission and higher antibody titres have been linked to lower rates of remission and worse kidney survival [16 -18 ].A 50% decrease in anti-PLA2R precedes a similar reduction in proteinuria by ≈10 months [18 ].
The length of the observation period should be individualized, but in general, initiation of IST is recommended if there is no trend towards improvement in disease activity ( decreasing proteinuria/anti-PLA2R titres, increasing serum albumin) after 6-9 months.In contrast, we strongly advise initiating immunosuppression without delay in patients with deteriorating kidney function not attributable to other causes ( e.g.diuretics overuse and high doses of RAAS blockers in the presence of low blood pressure) [19 , 20 ].Also, IST should be started in patients with massive oedema resistant to diuretics and/or severe complications of NS ( such as pulmonary thromboembolism) .Similarly, it seems reasonable not to wait in patients with massive proteinuria ( > 10-12 g/24 h) , severe hypoalbuminemia ( < 2 g/dl) and very high anti-PLA2R titres ( > 275 RU/ml) [18 ], as the probability of SR in these cases is low.

TIP 3. PRESCRIBE RITUXIMAB ( RTX) OR RTX COMBINED WITH CALCINEURIN INHIBITORS ( CNIS) IN PATIENTS CATEGORIZED AS MEDIUM RISK FOR PROGRESSIVE LOSS OF KIDNEY FUNCTION
We advise categorizing patients according to their risk factors for progressive loss of kidney function ( Fig. 1 ) and prescribing RTX or RTX combined with CNIs in medium-risk patients.Observational studies have shown a beneficial effect of monotherapy with RTX in MN [21 , 22 ] and this has been confirmed by several well-known prospective trials [23 -25 ] ( Table 1 ) .
Treatment with CNI ( tacrolimus, cyclosporine) has also proven to be effective [26 -30 ] ( Table 1 ) , but the major drawback of CNI monotherapy is the high rate of relapse after withdrawal.Risk factors for relapse include high residual proteinuria after partial remission ( PR) is attained and rapid withdrawal of the drug [28 ]; therefore, it is advisable to perform a very slow tapering ( 9-18 months, depending on the amount of residual proteinuria) [28 , 31 ] or to add RTX at the beginning of CNI withdrawal since the probability of relapse may be significantly reduced [32 , 33 ].
The proven efficacy of RTX and a better tolerance profile as compared with cyclophosphamide ( CYC) -based therapeutic regimens has led to the use of RTX as first-line therapy in most patients without severe risk factors.The ideal RTX dose is unclear ( Fig. 2 ) ; some studies have found that relatively low doses of RTX may result in worse outcomes than conventional doses [34 ].
One handicap of RTX is that the therapeutic effect is relatively slow and more than half of the patients persist with NS after 6 months.The latter can be countered by adding a CNI, with the advantage of a faster antiproteinuric effect, especially in cases with elevated proteinuria or severe hypoalbuminemia [33 , 35 ] ( Fig. 1 ) .More studies are needed to clarify the most effective regimens of RTX in monotherapy or combined with CNI.

TIP 4. PRESCRIBE CYC-BASED IMMUNOSUPPRESSION IN PATIENTS AT HIGH RISK FOR DISEASE PROGRESSION
The efficacy of alkylating agents alternating with glucocorticoids ( GCs) ( i.e. the modified Ponticelli regimen) has been demonstrated in several prospective studies [36 -40 ] and extended follow-up showed that alkylating agents decrease the risk of long-term kidney failure [41 ].Nevertheless, the use of CYC is currently restricted to high-risk patients ( Fig. 1 ) due to potential toxicity and the risk of serious adverse effects.Since CYC is associated with fewer side effects than chlorambucil [39 ], the latter has been almost completely relegated.
Few controlled studies have directly compared the Ponticelli regimen with other therapeutic alternatives [25 , 29 , 30 , 33 ], but CYC-based regimens have generally been demonstrated to induce a higher percentage of remissions ( Table 1 ) .A randomized controlled study demonstrated the superiority of cyclical chlorambucil plus GC over cyclosporine or supportive care in patients with declining kidney function [19 ].Likewise, observational studies have shown that CYC could be more effective than RTX in patients with high anti-PLA2R titres ( > 150 RU/ml) [42 ], although response to RTX in such cases has also been reported [43 ].Based on the above-mentioned data, we suggest a preferential use of CY-based regimens in high-risk patients ( Fig. 1 ) .The safety profile ( risk of sepsis, malignancy, myelotoxicity and infertility, among others) of CYC is closely related to the total cumulative dose [44 , 45 ].The modified Ponticelli regimen usually involves a cumulative dose of 10 g, but other oral CYC protocols may entail significantly higher doses [46 ].Different therapeutic CYC-based regimens have been proposed [47 -53 ] ( Fig. 2 ) .
The STARMEN trial ( NCT01955187) found that in the arm of cyclical CYC-GC, proteinuria had been significantly reduced and there was already a high rate of immunologic remissions after only 3 months of treatment ( cumulative CYC dose of 3.7 ± 1.1 g) ( Table 2 ) , suggesting that shorter courses may be as effective as the classically prescribed 6-month schedule.Also, recent studies advocate that the duration and cumulative dose of CYC can be reduced and individualized ac-cording to the patient's clinical and immunological response, either with cyclical oral CYC plus GC according to longitudinal anti-PLA2R measurements ( at 8, 16 and 24 weeks) [54 ] or with the use of pulse intravenous CYC instead of oral administration [50 , 52 , 53 ] ( Fig. 2 ) .Prospective studies comparing the modified Ponticelli regimen with lower-dose CYC schemes are needed.
A recent provocative retrospective case series described a high rate of complete remission ( CR) at 24 months with the combination of low-dose CYC plus RTX and GC [55 ].Combinations of CYC and RTX may be particularly attractive in high-risk patients-allowing a minimization of GC doses and a more rapid tapering-and deserve to be prospectively evaluated.

TIP 5. DO NOT USE GC MONOTHERAPY, MYCOPHENOLATE MOFETIL ( MMF) OR IMMUNOSUPPRESSANTS OTHER THAN RTX, CNI OR CYC IN THE INITIAL TREATMENT OF MN
Conflicting results were reported by two controlled trials performed several decades ago testing GC monotherapy in MN: deterioration of glomerular filtration rate was significantly more rapid in placebo-treated than in prednisone-treated patients in one study [56 ], while in a subsequent study [57 ], no significant differences were found between prednisone given on alternate days versus no specific treatment.Other observational works have also presented data in favour of and against GC monother-apy [58 , 59 ].In the more recent STARMEN trial, a significant decrease in proteinuria was observed ( Table 2 ) after the first month of GC therapy, even before starting the CYC cycle.In any case, given the inconsistent results between different studies, the known side effects of GC and the proven efficacy of other therapeutic strategies, the use of GC monotherapy for treating MN should be avoided.Although a potential beneficial effect of MMF in combination with GC, CNI or other immunosuppressants cannot be dismissed [60 , 61 ], MMF monotherapy did not show superiority as compared with conservative treatment in a prospective study [62 ].Currently, and until more conclusive data become available, the use of MMF in MN does not seem advisable.

TIP 6. DO NOT FORGET AVERTIBLE INFECTIOUS COMPLICATIONS IN PATIENTS RECEIVING IST
Patients with MN receiving immunosuppressive drugs are at increased risk for infections.Although specific recommendations on screening for latent infections and antimicrobial prophylaxis are currently lacking in subjects with MN, general recommendations for patients with glomerular diseases on immunosuppression have been outlined by international guidelines [63 ].It is worth noting that much of the evidence regarding this area is extrapolated from other settings, such as patients with rheumatic disease.
Latent tuberculosis infection should be ruled out according to local practice and treatment should be instituted along with IST in positive cases [63 ].Immunosuppressive agents ( particularly RTX) can induce hepatitis B virus reactivation/exacerbation [64 ], therefore, it is imperative to perform serologic tests ( HBsAg and HBcAb) in all patients and treat accordingly [65 ].Other pathogens that should be screened include hepatitis C virus, human immunodeficiency virus, syphilis and Strongyloides stercoralis ( only in high-risk populations) [63 , 66 ].
Pneumocystis prophylaxis should be strongly considered in patients receiving high-dose GC [67 ], RTX or CYC ( Table 3 ) .Current herpes zoster virus ( HZV) prophylaxis recommendations are vague [63 ].We advise the use of acyclovir or valacyclovir only in selected cases, such as highly immunosuppressed patients with repeated episodes of HZV or those on anti-CD38 antibodies.Hypogammaglobulinaemia should be monitored in RTX-treated patients since it has been associated with a significant increase in severe infections and increased mortality.Immunoglobulin replacement therapy may be of benefit in these cases [68 ].
Vaccination against pneumococcus ( heptavalent and 23valent) and influenza ( annually) should be performed in all patients.Prevention of HZV may be considered with the Shingrix vaccine.Recommendations regarding vaccination against severe acute respiratory syndrome coronavirus 2 are frequently amended, hence we advise following local or national guidelines, always considering that patients on IST fall into the highrisk category.

TIP 7. MONITOR AND PERSONALIZE IST BY CLOSELY MEASURING KIDNEY FUNCTION, PROTEINURIA, SERUM ALBUMIN AND ANTI-PLA2R LEVELS
In patients with anti-PLA2R positivity, following the tendency of antibody titres is highly useful as a guide to evaluate the efficacy of the chosen IST as well as dose and duration [12 , 13 , 17 , 18 , 69 , 70 , 71 ].The decrease in anti-PLA2R titres precedes a decrease in proteinuria by several weeks and an interval of 2.7 ± 1.7 months has been observed between immunologic and clinical remission [69 ].Patients initially considered resistant to RTX because of high baseline anti-PLA2R titres may reach remission after a second RTX course [72 ].
Although few centres routinely determine antibodies other than anti-PLA2R ( such as THSD7A, NELL-1 and others) , this scenario will likely change in the near future.Over the last few years, several antigens other than PLA2R with a clear pathogenic implication in NM have been identified [73 ].Their standardized determination will probably allow their monitoring in a similar manner to what happened with anti-PLA2R.On the other hand, the associations of these new antigens with different aetiologies and clinical manifestations ( autoimmune disorders, drugs, infections, tumours) will change our diagnostic approach, questioning the traditional separation between primary and secondary MN and offering a more personalized approach to diagnosis and treatment of the disease [73 ].
Monitoring of proteinuria, serum albumin and kidney function are also crucial, not just in anti-PLA2R-negative cases [74 ].Cases with immunologic remission not followed by remission of NS have been described [69 ] and recent evidence suggests that baseline anti-PLA2R titres do not predict disease progression with such confidence after adjusting for proteinuria and kidney function [75 , 76 ].A subanalysis of the MENTOR trial ( NCT01180036) showed that anti-PLA2R antibody titres combined with serum albumin levels after 3 months of treatment was the best method to evaluate the probability of remission [77 ].Serum albumin levels are of great value considering that proteinuria can show fluctuations caused by haemodynamic changes or inaccurate urine collection.Assessment of proteinuria should always be performed with the same method throughout the clinical course.A 24-h urine collection is the gold standard, but determination of the protein:creatinine ratio ( PCR) on an aliquot of an attempted 12-to 24-h urine collection is also acceptable.Avoid evaluation based on the albumin:creatinine ratio or random spot PCR [63 ].

TIP 8. TREATMENT OF RESISTANT DISEASE SHOULD BE PERFORMED IN A STEPWISE MANNER
Although there is no consensus on the definition of resistant MN, it is estimated that ≈30% of patients will not respond to treatment [78 ].If no immunologic ( persistence of high/unchanged anti-PLA2R antibodies) or clinical ( persistent NS/absence of at least PR) improvement is observed after 12 months following RTX or RTX plus CNI, resistant disease should be suspected.In these cases, CYC-based strategies are recommended as the next therapeutic step ( Fig. 3 ) , considering that the high efficacy of CYC in MN is supported by solid data [25 , 33 , 37 , 39 , 40 ] ( Table 1 ) .
Alternatively, new anti-CD20 monoclonal antibodies may be used in cases of non-response to RTX.Two small case series ( n = 13 patients) recently reported encouraging results with obinutuzumab in patients with MN refractory to RTX, achieving immunologic remission in most cases and CR or PR in up to 85% of patients [79 , 80 ].Positive results were also published for ofatumumab in RTX-intolerant or resistant patients ( n = 17) , whereas clinical response ( CR or PR) was observed in all cases and immunologic improvement in ≈50% [81 ].Obinutuzumab could be a good option in cases refractory or intolerant to RTX, whereas ofatumumab could work in cases with RTX intolerance or in those with anti-RTX antibodies.The latter agents should also be considered in patients without response to GC and CYC.
Non-response to anti-CD20 agents, CNI and CYC can represent a serious medical challenge.In these unfortunate cases, one may consider inclusion in clinical trials with experimental drugs or try anti-CD38 monoclonal antibodies such as daratumumab [82 ] or proteasome inhibition with bortezomib [83 ].

TIP 9. AFTER ATTAINING REMISSION, MONITOR RISK FACTORS FOR RELAPSE AND TREAT RELAPSES BASED ON THE PATIENT'S RISK PROFILE, SIMILAR TO THE FIRST BOUT OF THE DISEASE
The ideal outcome of treating MN with IST is to obtain CR.However, achieving PR is a satisfactory goal given that the risk of kidney failure is much lower than in patients without response [84 ].After PR is attained, residual proteinuria can be reduced by means of optimized non-immunosuppressive antiproteinuric treatments previously discussed ( tip 1) .
The prognosis of PR after IST is similar to that of spontaneous PR, yet a significant proportion of cases ( up to 47% in some studies) present with relapses [84 ].Relapses are more frequent in patients with PR compared with CR and in cases with persistent low serum albumin ( < 3.5 g/dl) [85 ].Persistence of mild hypoalbuminaemia despite non-nephrotic proteinuria is not uncommon and may indicate an active immunologic state.Beneficial effects of RTX have been described in patients with PR and persistent anti-PLA2R [86 ], although larger studies are needed.
In contrast, some patients may present with nephrotic-range proteinuria ( > 3.5 g/24 h) and normal serum albumin ( > 3.5 g/dl) .This subset of patients ( nephrotic-range proteinuria without NS) has been well characterized in FSGS [87 ] but seldom in MN.In our experience, this clinical profile can be observed in patients with PR in whom proteinuria increases to the nephrotic range, mainly due to significant weight gain.Optimization of RAAS blockade and other renoprotective drugs ( SGLT2i) together with weight loss usually decreases proteinuria to the non-nephrotic range.
Although there are no studies specifically focused on the treatment of relapses, it seems reasonable to select the type of IST based on risk factors ( Fig. 1 ) , similar to the first episode.However, if a new course of CYC-based therapies is considered, it is advisable that the cumulative dose does not exceed 10 g.

TIP 10. DO NOT FORGET THE UNDERLYING DISEASE THAT LED TO KIDNEY TRANSPLANTATION, AS MN MAY REOCCUR
Primary MN can relapse after kidney transplantation in ≈30-50% of cases [88 -90 ].Risk factors for recurrent disease include the presence of anti-PLA2R antibodies at the time of kidney transplantation ( especially if positivity persists during follow-up) , a weak transplant immunosuppressive regimen and an aggressive disease with rapid progression towards renal failure [88 , 91 , 92 ].
Proteinuria and immunologic monitoring should be performed frequently during the first 12-24 months after kidney transplantation.We suggest measuring 24-h proteinuria every 1-2 months during the first year and every 3 months subsequently, as well as anti-PLA2R determination at least every 3 months during the first year and every 4-6 months afterwards.An allograft biopsy should be indicated without delay in cases with increasing anti-PLA2R titres or worsening proteinuria ( Fig. 4 ) .
In contrast to native disease, clinical presentation can be highly variable ( positive histology without clinical expression, non-nephrotic proteinuria, NS) and spontaneous remissions are less frequently observed [93 , 94 ].Different studies have shown that MN recurrence is a risk factor for allograft failure [95 -97 ] and adjuvant IST may be required in some cases.After supportive therapy with antiproteinuric agents is initiated, close observation must be implemented.If proteinuria goes beyond 1 g/24 h, treatment with RTX ( Fig. 2 ) should be considered as first-line therapy [63 ].Reported remission rates with RTX are variable, but data from relatively large series suggest that partial or complete remission is observed in ≈70-80% of cases as compared with 30-35% with supportive therapy alone [88 , 98 ].Data on CYC-based regimens is quite scarce [89 ], but in case of non-response to RTX, its use could be considered.De novo MN ( unrelated to native kidney disease) is thought to be a type of alloimmune response since the majority of cases are associated with rejection lesions [99 -102 ], therefore therapy should be assessed on a case-by-case basis.

Figure 3 :
Figure 3: Treatment algorithm according to initial therapeutic regimen.

Figure 4 :
Figure 4: Algorithm for pretransplant risk recurrence of MN and post-transplant follow-up recommendations.

Table 3 : Antimicrobial prophylaxis for patients with MN on immunosuppressive treatment.
a Pneumocystis prophylaxis is based on trimethoprim-sulfamethoxazole [one double-strength ( 160/800 mg) tablet three times per week.Second-line treatments include dapsone 100 mg once daily, atovaquone 1500 mg once daily and pentamidine 300 mg ( nebulized) once monthly.